Question

Answer this question in a 1-2 paragraphs

The experiment that was performed was size exclusion chromatography. it involved:

- sephadex G75 (separates up to 80,000 g/mol)

- buffer: 20mM sodium Phosphate @ ph 7.0

- three compounds that were separated

1.) blue dextron (2,000,000 g/mol) blue

2.) cytochrome C (~12,000 g/mol) orange

3.) potassium chromate (194.2 g/mol) yellow

1 eluded first followed by 2 and then 3 as expected.

••• Come up with a creative, some sort of experiment that can possibly be performed to consolidate or build on the experiment already performed. “We are not looking for changes for the current experiment, this would be considered minor/ trivial changes and will lead to only 1 point”.

Answer 1

The compounds can be identified by simply measuring their absorbance at _max of each of these compounds. _max is the wavelength at which the absorbance of a compound is maximum and is characteristic of every compound. Blue Dextran has a _max of 620nm, cytochrome c being a protein will absorb maximally at 280nm & potassium chromate has a _max of around 330nm . The experiment consists of measuring the absorbance of each eluted fraction at each of these wavelengths and comparing them. The first fraction which consists of blue dextran should give maximal absorbance at 620nm followed by cytochrome c which should absorb maximally at 280nm. If each fraction is absorbing maximally at only one wavelength, this will ensure the purity of the fraction.

The compounds can be identified by injecting each fraction into a mass spectrometer, that ionises chemical species and sorts the ions based upon their mass to charge ratio. It is a highly sensitive technique that can even detect very low amount of the compond in a very accurate manner.